Mammalian organs are composed of cells surrounded by extracellular matrix (ECM) components. Interactions between cells and ECM initiate a flow of information that acts to regulate cell growth, cell migration, programmed cell death, and gene expression. These cell functions in turn affect the tissue integrity and destruction. The regulatory mechanisms by which ECM regulates cellular functions are under intensive study. Since ECM in intact tissue can influence cells in many ways: the biochemical interactions between ECM and their integrin receptors, the spatial arrangement of cell-cell interactions, the cell shape, the mechanical tension, and the cell dynamics, three-dimensional (3D) ECM lattices are developed to simulate the ECM-cell interaction in intact tissue. Cells embedded in this system demonstrate morphology and cellular functions usually observed under physiological condition. The regulation of matrix metalloproteinase I (MMP-1) expression is one of these functions. MMP-1 is principally responsible for collagen turnover in most tissues and, in particular, the skin. It has been found that cells grown in relaxed 3D collagen synthesized more MMP-1 than those in stressed 3D collagen. However, the regulation of MMP-1 expression in a tissue-like setting is far from clear. Taking a comparative approach that employs four types of 3D ECM lattices, relaxed collagen, stressed collagen, relaxed fibrin, and stressed fibrin, we were able to dissect mechanical and biochemical signals from 3D ECM. We demonstrated that relaxed, but not stressed 3D ECM (collagen or fibrin), regardless of biochemical nature, can induce MMP-1 expression, suggesting the mechanical nature of the regulation. However, the comparison between cells cultured in relaxed 3D collagen and fibrin showed the modulation of a different set of signaling molecules, suggesting that the physical signal transduction might employ different signaling pathways depending on the ECM environment. The objective of this grant proposal is to assess the mechanisms by which 3D ECM regulates cellular functions. We propose to use 3D collagen and fibrin as model systems to delineate the regulation of MMP-1, a functional output, in near-physiological condition. The mechanically relaxed and stressed 3D ECM lattices provide a physically distinct environment in the constant biochemical recognition between ECM-integrin receptors whereas the comparison between 3D collagen and fibrin lattices will shed light to the biochemical specificity in ECM mechanical signaling. Since 3D ECM is a tissue-equivalent system and MMP-1 is a gene product involved in embryogenesis, morphogenesis, and cancer development, as well as wound healing, understanding the signal transduction pathways from these ECM paradigms to the MMP-1 expression is of substantial importance.